The capacity of human secondary lymphoid tissue (LT) to act as a site for extrathymic de novo generation of T cells was investigated. A novel in vitro model for analysis of lymphopoiesis was developed using lymph node (LN) fragments conditioned with cytokines that support stem cell, stromal, and lymphoid growth and development. Following the addition of autologous CD34+ stem cells, pro ? and pre-T cells, as well as CD4CD8 double positive T cells were detected. The ability of LN tissue from HIV-infected subjects to support the development of these cellular populations, despite active in vitro viral replication, was demonstrated. The cellular and soluble factor parameters required for the development of lymphoid versus nonlymphoid populations were elucidated. The potential for the de novo generation of mature CD4+ or CD8+ T cells with a diverse TCR repertoire is being further investigated. - HIV; lymphopoiesis; lymphoid tissue; thymus; cytokines; CD34+ stem cells; T cells